U.S. patent application number 11/650567 was filed with the patent office on 2007-07-19 for quinaldine based semisquaraines and squaraine dyes, process for preparation thereof and use thereof.
Invention is credited to Kalliat Thazhathveetil Arun, Jyothish Kuthanapillil, Danaboyina Ramaiah.
Application Number | 20070167350 11/650567 |
Document ID | / |
Family ID | 38228594 |
Filed Date | 2007-07-19 |
United States Patent
Application |
20070167350 |
Kind Code |
A1 |
Ramaiah; Danaboyina ; et
al. |
July 19, 2007 |
Quinaldine based semisquaraines and squaraine dyes, process for
preparation thereof and use thereof
Abstract
The present invention provides quinaldine-based semisquaraines,
symmetrical and unsymmetrical squaraine dyes represented by the
general formulae 1, 2 and 3 and/or pharmaceutically acceptable
derivatives thereof as sensitizers for photodynamic therapeutical
and industrial applications. These symmetrical and unsymmetrical
squaraine dyes posses absorption which extends well into the
photodynamic window (650-800 nm) and hence are useful for the
treatment of deep seated tumors. The absorption of these dyes can
be tuned by changing the substituents on the quinaldine moiety
thereby enabling the development of a library of dyes which have
absorption ranging from 650 to 800 nm. They also exhibited
fluorescence emission in the long wavelength region making them
useful as near infrared fluorescence sensors for the detection of
tumors. These dyes are non-toxic in the dark and exhibit good
photocytotoxicity. Accordingly the quinaldine based semisquaraines,
symmetrical and unsymmetrical squaraines are extremely useful as
diagnostic and therapeutic agents for photodynamic therapeutical
and industrial applications.
Inventors: |
Ramaiah; Danaboyina;
(Kerala, IN) ; Kuthanapillil; Jyothish; (Kerala,
IN) ; Arun; Kalliat Thazhathveetil; (Kerala,
IN) |
Correspondence
Address: |
NIXON & VANDERHYE, PC
901 NORTH GLEBE ROAD, 11TH FLOOR
ARLINGTON
VA
22203
US
|
Family ID: |
38228594 |
Appl. No.: |
11/650567 |
Filed: |
January 8, 2007 |
Current U.S.
Class: |
514/23 ; 514/176;
514/311; 530/300; 536/17.4; 540/107; 546/176 |
Current CPC
Class: |
C07D 215/14 20130101;
A61P 35/00 20180101; C09B 57/007 20130101 |
Class at
Publication: |
514/002 ;
514/023; 514/176; 514/311; 530/300; 536/017.4; 540/107;
546/176 |
International
Class: |
A61K 38/16 20060101
A61K038/16; A61K 31/7052 20060101 A61K031/7052; C07J 43/00 20060101
C07J043/00; C07D 215/12 20060101 C07D215/12; C07K 14/47 20060101
C07K014/47; C07H 17/02 20060101 C07H017/02 |
Foreign Application Data
Date |
Code |
Application Number |
Jan 6, 2006 |
IN |
60/DEL/2006 |
Claims
1. A semisquaraine of formula 1 and pharmaceutically acceptable
derivatives thereof ##STR7## wherein, X=H, I, Br, NO.sub.2, CN,
COOH, SO.sub.3H or OR.sup.1, wherein R.sup.1=H, alkyl groups having
from 1 to 6 carbon atoms, glycols having from 2 to 4 ethylene
groups, cholic acid, cholesterol, sugar, amino acids or peptides,
wherein Y=H, I, or Br, wherein, Z=H, I, Br, NO.sub.2, CN, COOH,
SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein R.sup.2=alkyl groups
having from 1 to 6 carbon atoms, glycols having from 2 to 4
ethylene groups, alkyl sulphonic acids and acid chlorides having
from 1 to 6 carbon atoms.
2. A process for the preparation of a semisquaraine of the formula
1 ##STR8## wherein, X=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H or
OR.sup.1, wherein R.sup.1=H, alkyl groups having from 1 to 6 carbon
atoms, glycols having from 2 to 4 ethylene groups, cholic acid,
cholesterol, sugar, amino acids or peptides, wherein Y=H, I, or Br,
wherein, Z=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or
OR.sup.1, wherein R.sup.2=alkyl groups having from 1 to 6 carbon
atoms, glycols having from 2 to 4 ethylene groups, alkyl sulphonic
acids and acid chlorides having from 1 to 6 carbon atoms, said
process comprising reacting quinaldinium salt and squaric acid in a
mixture of benzene and n-butanol under stirring, removing the
solvent and purifying the residue obtained to obtain compound of
formula 1.
3. A process as claimed in claim 2 wherein the quinaldinium salt
and squaric acid are taken in a ratio of 1:1.
4. A process as claimed in claim 2 wherein the mixture of benzene
and n-butanol is 1:1 mixture.
5. A process as claimed in claim 2 wherein the stirring is carried
out at a temperature in the range of 90-110.degree. C. and for a
time period of 24-36 h.
6. A process as claimed in claim 2 wherein the solvent is removed
by evaporation.
7. A process as claimed in claim 2 wherein the purification is
effected by column chromatography over silica gel.
8. Quinaldine based symmetrical squaraine dye of formula 2 below or
pharmaceutically acceptable salts thereof ##STR9## wherein, X=H, I,
Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein,
R.sup.1=H, alkyl groups having from 1 to 6 carbon atoms, glycols
having from 2 to 4 ethylene groups, cholic acid, cholesterol,
sugar, amino acids or peptides, wherein, Y=H, I, Br, NO.sub.2, CN,
COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein R.sup.2=alkyl
groups having from 1 to 6 carbon atoms, glycols having from 2 to 4
ethylene groups, alkyl sulphonic acids and acid chlorides having
from 1 to 6 carbon atoms.
9. Process for the preparation of a quinalidine based symmetrical
squaraine dye of formula 2 ##STR10## wherein, X=H, I, Br, NO.sub.2,
CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein, R.sup.1=H,
alkyl groups having from 1 to 6 carbon atoms, glycols having from 2
to 4 ethylene groups, cholic acid, cholesterol, sugar, amino acids
or peptides, wherein, Y=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H,
SO.sub.2Cl or OR.sup.1, wherein R.sup.2=alkyl groups having from 1
to 6 carbon atoms, glycols having from 2 to 4 ethylene groups,
alkyl sulphonic acids and acid chlorides having from 1 to 6 carbon
atoms, said process comprising reacting a quinaldinium salt and
squaric acid in a solution of benzene and n-butanol with stirring,
removing said solvent and then purifying the residue so obtained to
obtain the squaraine dye of formula 2.
10. A process as claimed in claim 9 wherein the quinaldinium salt
and squaric acid are taken in a ratio of 2:1.
11. A process as claimed in claim 9 wherein the mixture of benzene
and n-butanol is 1:1 mixture.
12. A process as claimed in claim 9 wherein the stirring is carried
out at a temperature in the range of 90-110.degree. C. and for a
time period of 24-36 h.
13. A process as claimed in claim 9 wherein the solvent is removed
by evaporation.
14. A process as claimed in claim 9 wherein the purification is
effected by column chromatography over silica gel.
15. A quinaldine based asymmetrical squaraine dye of formula 3 and
pharmaceutically acceptable derivatives thereof ##STR11## wherein,
X=OR.sup.1, wherein, R.sup.1=H, alkyl groups having from 1 to 6
carbon atoms, glycols having from 2 to 4 ethylene groups, cholic
acid, cholesterol, sugar, amino acids or peptides, wherein Y=H, I,
Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein
Q=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein Z=H,
I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein
R.sup.2=alkyl groups having from 1 to 6 carbon atoms, glycols
having from 2 to 4 ethylene groups, alkyl sulphonic acids and acid
chlorides having from 1 to 6 carbon atoms.
16. A process for the preparation of a quinaldine based
asymmetrical squaraine dye of formula 3 ##STR12## wherein,
X=OR.sup.1, wherein, R.sup.1=H, alkyl groups having from 1 to 6
carbon atoms, glycols having from 2 to 4 ethylene groups, cholic
acid, cholesterol, sugar, amino acids or peptides, wherein Y=H, I,
Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein
Q=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein Z=H,
I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein
R.sup.2=alkyl groups having from 1 to 6 carbon atoms, glycols
having from 2 to 4 ethylene groups, alkyl sulphonic acids and acid
chlorides having from 1 to 6 carbon atoms, said process comprising
heating a quinaldine based semisquaraine of formula 1 and
quinaldinium salts substituted with halogen atoms at 6-position in
a mixture of benzene and n-butanol, removing the solvent to obtain
a residue which on purification provides compound of formula 3.
17. A process as claimed in claim 16 wherein the mixture of benzene
and n-butanol is a 1:1 mixture.
18. A process as claimed in claim 16 wherein the heating is carried
out to a temperature in the range of 90-110.degree. C. and for a
time period of 12-18 h.
19. A process as claimed in claim 16 wherein the solvent is removed
by evaporation.
20. A process as claimed in claim 16 wherein the purification is
effected by column chromatography over silica gel.
21. Use of compound of formula 1 as a photosensitizer in
photodynamic treatment of cancer and other related diseases.
22. Use of compound of formula 1 as a near infrared fluorescent
sensor for diagnosis of cancer.
23. Use of compound of formula 1 as a sensitizer in sterilization
of fluids such as water.
24. Use of compound of formula 2 as a photosensitizer in
photodynamic treatment of cancer and other related diseases.
25. Use of compound of formula 2 as a near infrared fluorescent
sensor for diagnosis of cancer.
26. Use of compound of formula 2 as a sensitizer in sterilization
of fluids such as water.
27. Use of compound of formula 3 as a photosensitizer in
photodynamic treatment of cancer and other related diseases.
28. Use of compound of formula 3 as a near infrared fluorescent
sensor for diagnosis of cancer.
29. Use of compound of formula 3 as a sensitizer in sterilization
of fluids such as water.
Description
[0001] The present invention relates to quinaldine based
semisquaraines of the general formula 1 and squaraine dyes of the
general formulae 2 and 3, respectively as shown below ##STR1##
wherein, X=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H or OR.sup.1,
wherein R.sup.1=H, alkyl groups having from 1 to 6 carbon atoms,
glycols having from 2 to 4 ethylene groups, cholic acid,
cholesterol, sugar, amino acids or peptides, wherein Y=H, I, or Br,
wherein, Z=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or
OR.sup.1, wherein R=alkyl groups having from 1 to 6 carbon atoms,
glycols having from 2 to 4 ethylene groups, alkyl sulphonic acids
and acid chlorides having from 1 to 6 carbon atoms ##STR2##
wherein, X=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or
OR.sup.1, wherein, R.sup.1=H, alkyl groups having from 1 to 6
carbon atoms, glycols having from 2 to 4 ethylene groups, cholic
acid, cholesterol, sugar, amino acids or peptides, wherein, Y=H, I,
Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein
R.sup.2=alkyl groups having from 1 to 6 carbon atoms, glycols
having from 2 to 4 ethylene groups, alkyl sulphonic acids and acid
chlorides having from 1 to 6 carbon atoms ##STR3## wherein,
X=OR.sup.1, wherein, R.sup.1=H, alkyl groups having from 1 to 6
carbon atoms, glycols having from 2 to 4 ethylene groups, cholic
acid, cholesterol, sugar, amino acids or peptides, wherein Y=H, I,
Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein
Q=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein Z=H,
I, Br, NO.sub.2, CN, COOH, SO.sub.3H, SO.sub.2Cl, wherein
R.sup.2=alkyl groups having from 1 to 6 carbon atoms, glycols
having from 2 to 4 ethylene groups, alkyl sulphonic acids and acid
chlorides having from 1 to 6 carbon atoms; and pharmaceutically
acceptable derivatives thereof for photodynamic therapeutical,
diagnostic and industrial applications.
[0002] The present invention also relates to a process for the
preparation of quinaldine based semisquaraines of the general
formula 1 and squaraine dyes of the general formulae 2 and 3 and
use of such sensitizers for photodynamic, therapeutical, diagnostic
and industrial applications.
[0003] The present invention also relates to semisquaraines and
squaraine dyes of the general formulae 1, 2 and 3 or
pharmaceutically acceptable derivatives thereof, for use as
photosensitizers in photodynamic applications for the detection and
treatment of cancer and other diseases in human beings or
animals.
[0004] The present invention also relates to semisquaraines and
squaraine dyes of the general formulae 1, 2 and 3 or
pharmaceutically acceptable derivatives thereof for use as
near-infrared fluorescence probes for biological applications.
[0005] The present investigation also relates to quinaldine based
semisquaraines of the general formula 1, which can be used as
potential protein tyrosine phosphatases (PTPases) inhibitors and
squaraine dyes of the general formulae 2 and 3 for use in the
detection of metal ions in the biological media.
[0006] The present invention also relates to a process for the
preparation of quinaldine based semisquaraines of the general
formula 1 and squaraine dyes of the general formulae 2 and 3 and/or
their derivatives for photodynamic industrial applications such as
sterilization of fluids and water and related other
applications.
[0007] Photodynamic therapy is an emerging modality for the
diagnosis and treatment of cancer and various diseases. The large
body of evidence suggests that photodynamic therapy represents a
convenient and effective approach for a variety of cancers. The
process requires the presence of a photosensitizing agent, which is
capable of being taken up by target cells and tissues and which, on
irradiation by light of a particular wavelength, generates species
which are toxic to those cells and tissues. Photodynamic therapy
has advantages over many other conventional therapies due to the
selectivity of the photodynamic process. There is more sensitizer
in tumor tissues than in the normal tissues. This reduces the
potential for destruction of normal tissues. In addition the
ability to direct light specifically onto the target cells and
tissues by the use of fiber-optic technology further increased the
selectivity of this process. Furthermore, the use of
photosensitizing agents, which produce no response until irradiated
with light, reduces significantly the potential for side effects,
which may complicate the process.
[0008] The only sensitizer that has been extensively studied is a
hematoporphyrin derivative (HpD), also known as first generation
photosensitizer. HpD and its commercial variants Photofrin (porfine
sodium), Photosan, Photogen were the first ones to be approved in
clinical use and for which first regulatory authorizations were
obtained. However, Photofrin is at the disadvantage of being a
mixture of products the composition of which is highly sensitive to
the synthetic methodology adopted. It is known to cause cutaneous
photosensitivity as an undesirable side effect because of its slow
release from the body. Under these circumstances, a patient treated
with Photofrin is required to stay in the dark for a long period
until it is excreted from the body. Photofrin possesses only weak
absorption in the red region of spectrum (the molar absorption
coefficient being as small as 3000 M.sup.-1 cm.sup.-1 at 630 nm),
leading to difficulty in delivering light to some tumor sites and
also incomplete light penetration of larger tumors. Therefore,
photodynamic therapy using Photofrin is only indicated for cancers
developing in the surface layers of less than 10 mm depth.
References may be made to Dougherty, T. J. Photochem. Photobiol.
1987, 45, 879; Kessel, D.; Dougherty, T. J. Phorphyrin
Photosensitization; Plenum Publishing Corp. New York, 1983; Brown,
S. B.; Truscott, T. G. Chem. Ber, 1993, 29, 955; Andreoni, A.,
Cubeddu, R. Phorphyrins in Tumor Phototherapy; Plenum Publishing
Corp.: New York, 1984; Brasseur, N.; Hasarat, A., Langlois, R.,
Wagner, J. R; Roussean, J.; van Lier J. E. Photochem. Photobiol.
1987, 45, 581; Spikes, J. D, Photochem. Photobiol. 1986, 43, 691;
Firey, P. A.; Ford, W. E.; Sounik, J. R.; Kenney, M. E.; Rodgers,
M. A. J. J. Am. Chem. Soc. 1988, 110, 7626; Moan, J. Cancer Lett.
1986, 33, 45; Tralau, C J., Young, A. R.; Walker, N. P. J.; Vernon,
D. I.; MacRobert, A. J., Brown, S. B.; Brown, S. G. Photochem.
Photobiol. 1989, 49, 305. Bonnett, R. Chem. Soc. Rev. 1995, 24, 19;
Lane, N. Scientific American 2003, 38-45.
[0009] To overcome the drawbacks of the first generation
sensitizers, second generation photosensitizers that exhibit strong
absorption in the long wavelength region have been synthesized.
Second generation sensitizers that are under evaluation at various
clinical phases of photodynamic therapy include chlorins,
porphycenes, benzoporphyrins, phthalocyanins, purpurins and
aminolevulinic acid-mediated porphyrins. Purpurins possess
favorable optical properties and biodistribution patterns but
require solubilizing or emulsifying agents such as liposomes or
lipoproteins for their photodynamic applications. Chlorins have
strong absorption in the red and infrared regions of the spectrum
and compete favorably with Photofrin, but skin photosensitivity is
a major problem with them. Phthalocyanins and metallophthalocyanins
have been found to have strong absorption in the 600-700 nm
regions, but details of the extent of sulfonation versus the
photodynamic activity is not clear.
[0010] References may be made to U.S. Pat. Nos. 603,267; 5,965,598;
5,889,181; 586,035; 5,789,586; Kostenich, G. A.; Zuravkin, I. N.;
Zhavrid, E. A. J Photochem. Photobiol. B. Biol. 1994, 22, 211;
Leach, M. W.; Higgins, R. J.; Autry, S. A.; Boggan, J. E.; Lee,
S.-J. H.; Smith, K. M. Photochem. Photobiol. 1993, 1616, 275;
Vogel, E.; Kocher, M Schmickler, H.; Lex, J. Angew. Chem. Int. Ed.
Engl. 1986, 25, 197; Leunig, M.; Richert, C.; Gamarra, F.; Lumper,
W.; Vogel, E.; Jochani D.; Goetz, A. E. Br. J. Cancer 1993, 68,
225; Boyle, R. W.; Legnoff, C. C.; Vanheir, J. E. Br. J. Cancer,
1993, 67, 1177; Wohrl, D.; Shopova, M.; Muller, S.; Muleiv, A. D.;
Mantereva, V. N.; Krastev, K. K. J Photochem. Photobiol. B. Biol.
1993, 21, 155; Morgan, A. R.; Garbo, G. M.; Keck, R. W.; Ericksen,
L. D.; Selman, S. H. Photochem. Photobiol. 1990, 51, 589.
[0011] Development of photosensitizers, which have strong
absorptions in the long wavelength region, non-toxic to normal
tissues, soluble in buffer at physiological pH, can be bleached
during the photodynamic treatment and exhibit higher therapeutic
efficacy are still desired. Also the design of functional molecules
that can target specific cancer cells are extremely important
because of the biochemical and biomedical applications.
[0012] Squaraines form a class of dyes possessing sharp and intense
absorption bands in the red to near infra red region. The
photophysical and photochemical properties of these have been
studied extensively, because their absorption and photochemical
characteristics make them highly suitable for a number of
industrial applications. These include photoreceptors in copiers,
photoconductors in organic solar cells and IR absorbers in organic
optical disks. Squaraines find industrial applications in
xerographic photoreceptors, solar cells and optical recording
devices. However, due to the very low intersystem crossing
efficiency of these dyes, their potential as photosensitizers in
photodynamic therapeutical applications has not yet been explored.
References may be made to U.S. Pat. Nos. 6,001,523; 5,552,253;
5,444,463; Law, K.-Y. Chem. Rev. 1993, 93, 449; Piechowski, A P;
Bird, G. R.; Morel, D L.; Stogryn, E. L. J. Phy. Chem. 1984, 88,
934. Accordingly, the use of squaraine based dyes was studied to
observe if problems associated with the prior art could be
overcome. Preliminary investigations indicated that halogenation of
the squaraine moiety resulted in increased water solubility and
intersystem crossing efficiency when compared to the parent
unsubstituted squaraine dyes. These halogenated dyes exhibited
strong absorption in the near infrared region (>600 nm) and
significant bathochromic shifts in presence of microheterogeneous
media. Triplet excited states were the main transients involved in
these systems, which interact efficiently with molecular oxygen
generating biologically highly reactive singlet oxygen in
quantitative yields thereby making them potential candidates in
photodynamic therapeutical applications. We have also investigated
the cytotoxicity and mutagenicity of these dyes in the dark and
under irradiation conditions. The results demonstrate that the
halogenated squaraine dyes exhibit efficient cytotoxicity upon
photoexcitation in two different types of mammalian cell lines
(AS52 Chinese hamster ovary cells and L5178Y mouse limphoma cells).
The cytotoxicities of the compounds under dark are much lower.
Mechanistically, the cellular damage may be mediated by singlet
oxygen, generated from photoexcited squaraine dyes. The squaraine
dyes are also found to be only weekly mutagenic in the presence and
absence of light. References may be made to Ramaiah, D.; Arun, K.
T.; Das, S. and Epe, B. U.S. Pat. No. 6,770,787B2 (2004), Ramaiah,
D.; Arun, K. T.; Das, S. and Epe, B. Indian patent No. 193540
(2004). Ramaiah, D.; Joy, A.; Chandrasekhar, N; Eldho, N. V.; Das,
S.; George, M. V. Photochem. Photobiol. 1997, 65, 783; Arun, K. T.;
Ramaiah, D.; Epe, B. J. Phys. Chem. B 2002, 107, 11622, Ramaiah,
D.; Eckert, I; Arun, K. T.; Weidenfeller, L.; Epe, B. Photochem.
Photobiol. 2002, 76, 672; Ramaiah, D.; Eckert, I; Arun, K. T.;
Weidenfeller, L.; Epe, B. Photochem. Photobiol. 2004, 79, 99.
[0013] However, the absorption of these dyes is in the lower part
of the photodynamic window, .about.600 nm where most of the
biological tissues absorb light. Hence the use of these compounds
for the destruction of deep-seated tumors becomes difficult. In
this context the present invention aims at the development of
efficient sensitizers based on squaraine dyes which can be used for
tumors located deep inside the body. Quinaldine based squaraine
dyes are known to have absorption well beyond 700 nm, which suits
well for applications in photodynamic therapy. References may be
made to Santos, P. F.; Reis, L. V.; Almeida, P.; Oliveira, A. S.;
Ferreira, L. F. V. J. Photochem. Photobiol. A: Chem. 2003, 160,
159.
[0014] The present invention is an attempt to develop
squaraine-based dyes having absorption in the near infrared region,
which can be used for the destruction as well as detection of
deep-seated tumors. We have substituted the quinaldine based
squaraine dyes with heavy atoms like bromine and iodine which
enhance their intersystem crossing efficiency and thereby increase
their ability to generate cytotoxic agents like singlet oxygen. We
have also introduced various substituents on the benzo ring of the
quinaldine moiety, which enables their physiological uptake.
Moreover, we have developed conjugates of dye and cellular
recognition elements like sugar, proteins, cholic acid and
cholesterol, which enables site specific destruction of tumors.
References may be made to Monsigny, M.; Roche, A. C.; Midoux, P.;
Kieda, C.; Mayer, R. In Lectins and Glycoconjugates in Oncology:
Structure, Function, Clinical Application; Gabius, H. J.; Nagel, G.
A. Eds.; Springer: Heidelberg, 1988, p 1.
[0015] In the present invention we have synthesized novel
quinaldine based squaraine dyes and investigated their
photophysical properties under different conditions and
photobiological properties in mammalian cell lines like mouse
lymphoma L1210 cells.
[0016] The main object of the present invention is to provide
efficient squaraine based dyes and/or pharmaceutical acceptable
derivatives thereof, for use as sensitizers in photodynamic
therapeutical applications including the treatment of cancer.
[0017] Another object of the invention is to provide efficient
squaraine based dyes and/or pharmaceutical acceptable derivatives
thereof, for use as sensitizers in photodynamic therapeutical
applications for the detection of tumors.
[0018] Another object of the invention is to provide efficient
squaraine based dyes and/or pharmaceutical acceptable derivatives
thereof, for use as near-infrared fluorescence sensors for
biological applications.
[0019] Still another object of the invention is to provide cellular
recognition elements linked squaraine dyes like sugar, cholesterol
and amino acids linked squaraine dyes and acceptable derivatives
thereof in improving their efficiency as sensitizers for
photodynamic applications.
[0020] Yet another object of the invention is to provide an
efficient method for the synthesis of quinaldine based
unsymmetrical squaraine dyes and derivatives thereof.
[0021] Yet another object of the invention is to provide an
efficient method for the synthesis of quinaldine-based
semisquaraines and derivatives thereof.
[0022] Yet another object of the invention is to provide quinaldine
based semisquaraines, which can be used as potential protein
tyrosine phosphatases (PTPases) inhibitors.
[0023] Another object of the invention is to provide squaraine
based dyes and/or their derivatives for use as photodynamic
industrial applications such as sterilization of fluids, water and
related other applications.
[0024] Yet another object of the invention is to provide quinaldine
based symmetrical and unsymmetrical squaraine dyes and
semisquaraines and/or their derivatives thereof that can be used
for the detection of biologically important metal ions under
physiological condition.
[0025] In the drawings accompanying the specifications:
[0026] FIG. 1: Absorption and fluorescence emission spectra of
semisquaraine of general formula 1, where X=OH, Y=H, Z=H,
R.sup.2=CH.sub.3, in methanol
[0027] FIG. 2: Absorption and fluorescence emission spectra of
squaraine dyes of general formula 2, where X=I, Y=H,
R.sup.2=CH.sub.3, in dimethyl sulfoxide.
[0028] FIG. 3: Change in absorption spectra obtained for the
reaction between the semisquaraine of the general formula 1, where
X=OH, Y=H, Z=H, R.sup.2=CH.sub.3 and the 6-iodoquinaldinium salt at
various time intervals. Inset shows the change in absorption
spectra obtained for the reaction between the semisquaraine of the
general formula 1, where X=OH, Y=H, Z=H, R.sup.2=CH.sub.3 and the
6-hydroxyquinaldinium salt at various time intervals.
[0029] FIG. 4: Absorption and fluorescence emission spectra of
unsymmetrical squaraine dyes of general formula 3, where
X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3, in
dimethyl sulfoxide.
[0030] FIG. 5: Photocytotoxicity of the squaraine dyes of general
formula 2, where X=I, Y=H, R.sup.2=CH.sub.3, and general formula 3,
where X=OH, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3 in L1210 mouse lymphoma
cells. Data gives the number of cell (% of controls) counted after
48 h after the treatment for 22.5 min at 0.degree. C. with various
concentrations of the squaraine dyes under dark and under
irradiation. Data points represent the mean of three independent
experiments (.+-.SD).
[0031] FIG. 6: Photocytotoxicity of the sugar linked and
cholesterol-squaraine dyes of general formula 3, where
X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3 and where
X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3 L1210
mouse lymphoma cells. Data gives the number of cell (% of controls)
counted after 48 h after the treatment for 22.5 min at 0.degree. C.
with various concentrations of the squaraine derivatives under dark
and under irradiation. Data points represent the mean of three
independent experiments (.+-.SD).
[0032] Accordingly, the present invention relates to quinaldine
based squaraine dyes of the general formulae 2 and 3 and
semisquaraines of the general formula 1 and pharmaceutically
acceptable derivatives thereof. ##STR4## wherein, X=H, I, Br,
NO.sub.2, CN, COOH, SO.sub.3H or OR.sup.1, wherein R.sup.1=H, alkyl
groups having from 1 to 6 carbon atoms, glycols having from 2 to 4
ethylene groups, cholic acid, cholesterol, sugar, amino acids or
peptides, wherein Y=H, I, or Br, wherein, Z=H, I, Br, NO.sub.2, CN,
COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein R.sup.2=alkyl
groups having from 1 to 6 carbon atoms, glycols having from 2 to 4
ethylene groups, alkyl sulphonic acids and acid chlorides having
from 1 to 6 carbon atoms ##STR5## wherein, X=H, I, Br, NO.sub.2,
CN, COOH, SO.sub.3H, SO.sub.2Cl or OR.sup.1, wherein, R.sup.1=H,
alkyl groups having from 1 to 6 carbon atoms, glycols having from 2
to 4 ethylene groups, cholic acid, cholesterol, sugar, amino acids
or peptides, wherein, Y=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H,
SO.sub.2Cl or OR.sup.1, wherein R.sup.2=alkyl groups having from 1
to 6 carbon atoms, glycols having from 2 to 4 ethylene groups,
alkyl sulphonic acids and acid chlorides having from 1 to 6 carbon
atoms ##STR6## wherein, X=OR.sup.1, wherein, R.sup.1=H, alkyl
groups having from 1 to 6 carbon atoms, glycols having from 2 to 4
ethylene groups, cholic acid, cholesterol, sugar, amino acids or
peptides, wherein Y=H, I, Br, NO.sub.2, CN, COOH, SO.sub.3H,
SO.sub.2Cl or OR.sup.1, wherein Q=H, I, Br, NO.sub.2, CN, COOH,
SO.sub.3H, SO.sub.2Cl, wherein Z=H, I, Br, NO.sub.2, CN, COOH,
SO.sub.3H, SO.sub.2Cl, wherein R.sup.2=alkyl groups having from 1
to 6 carbon atoms, glycols having from 2 to 4 ethylene groups,
alkyl sulphonic acids and acid chlorides having from 1 to 6 carbon
atoms.
[0033] In one embodiment of the invention, the quinaldinium salt
and squaric acid in the ratio 1:1 in a mixture of benzene and
n-butanol (1:1) is refluxed at (90-110.degree. C.) for a time
period of 24-36 h. Removal of the solvent gave a residue, which was
then subjected to column chromatography over silica gel to obtain
compounds of the general formula 1.
[0034] In another embodiment of the of the invention, the
quinaldinium salt and squaric acid in the ratio (2:1) in a solution
of benzene and n-butanol (1:1) is refluxed for 24-36 h. Removal of
the solvent gave a residue, which was then subjected to column
chromatography over silica gel to obtain compounds of the general
formula 2.
[0035] In another embodiment of the invention, the quinaldine based
semisquaraines of the general formula 1 and the quinaldinium salts
substituted with heavy atoms (halogen atoms) at the 6-position are
heated in a mixture of benzene and n-butanol (1:1) at
(90-110.degree. C.) for a time period of 12-18 h. Removal of the
solvent gave a residue, which was then subjected to column
chromatography over silica gel to obtain compounds of the general
formula 3.
[0036] In another embodiment of the invention, the compound of the
formulae 1, 2 and 3 are used as photosensitizers in the
photodynamic treatment of cancer and other related diseases.
[0037] In yet another embodiment of the invention, the compounds of
the formulae 1, 2 and 3 are used as near infrared fluorescent
sensors for the diagnosis of cancer.
[0038] The invention also relates to the use of compound of
formulae 1, 2 and 3 as a sensitizer in the sterilization of fluids,
water and related other applications.
[0039] In the present investigation, the quinaldine based
semisquaraines of the general formula 1 have been synthesized.
These semisquaraines were found to be unreactive towards
quinaldinium salts containing electron-donating substituents.
However, when refluxed with quinaldinium salts containing
electron-withdrawing substituents like nitro, cyano and
electro-negative substituents like halogen atoms, it is observed
that the semisquaraines of the general formula 1 formed squaraine
dyes of the general formula 3. Thus a highly efficient, two-step
process for the synthesis of unsymmetrical squaraine dyes has been
demonstrated.
[0040] In the present invention, novel quinaldine based squaraine
dyes were synthesized and their photophysical and in vitro
photobiological properties in the presence and absence of light
were investigated. These dyes were found to have absorption in the
near infrared region (700-750 nm). In the preparation of the
compounds of the general formulae 2 and 3, the aromatic ring of the
quinaldine moiety is modified with heavier halogen atoms such as
bromine and iodine, this is to enable intersystem crossing
efficiency of these dyes, which, in turn, would enhance the
generation of highly reactive species like singlet oxygen.
[0041] In the present investigation, the aromatic ring of the
quinaldine moiety is yet again modified with cholesterol and sugar
moieties. Modification with cholesterol and sugar moieties is
expected to increase the cell permeability and to bring about
target specificity to these dyes since cholesterol is known to have
membrane affinity and an accumulation of the esters in lipoprotein
particles--and in consequence in proliferating cells and sugars are
known to have affinity to the cancer cells.
[0042] In the present invention, we have investigated the
cytotoxicity of these dyes in mammalian AS52 cells both in the
presence and absence of light. It was found that the cytotoxicity
of (sugar linked) squaraine dyes of general formula 3, where
X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3 and
(cholesterol linked) where X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I,
Z=H, R.sup.2=CH.sub.3 are significantly high when compared with the
(hydroxyl-substituted derivative) squaraine dye of general formula
3, where X=OH, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3. These dyes were
found to be more cytotoxic than the diiodo squaraine dye of the
general formula 2, where X=I, Y=H, R.sup.2=CH.sub.3, which may be
attributed to their increased cell permeability due to the presence
of sugar and cholesterol moieties.
[0043] The following examples are given by way of illustration and
therefore should not be construed to limit the scope of present
investigation.
[0044] Examples 1-3 represent typical synthesis of compounds of the
general formulae 1-3 and examples 4-7 represent the photophysical
properties of general formulae 1-3 and examples 8 and 9 represent
the in vitro cytotoxic properties of quinaldine based squaraine
dyes of formulae 2 and 3.
EXAMPLE 1
General Procedure for the Preparation of Compounds Represented by
Formula 1
[0045] A mixture of the corresponding quinaldinium salt (1 mmol),
squaric acid (1 mmol) and quinoline (0.5 mL) was refluxed in a
mixture of n-butanol and benzene (6-9 mL each, 1:1) with azeotropic
distillation of water for 24-36 h. The solvent was distilled off
under reduced pressure to obtain a residue which was
chromatographed over silica gel. Elution of the column with a
mixture (1:19) of methanol and chloroform gave the semisquaraines
of the general formula 1.
[0046] The physiochemical properties of compound of the general
formula 1, (wherein X=OH, Y=H and Z=H, R.sup.2=CH.sub.3, 90-95%),
mp 150-152.degree. C., IR (KBr) .nu..sub.max 3412, 3042, 2963,
1761, 1606 cm.sup.-1. .sup.1H-NMR (CDCl.sub.3+DMSO-d.sub.6, 1:4)
.delta. 10.06 (1H, OH), 9.29 (1H, d, J=11.4 Hz), 7.89 (1H, d, J=9.3
Hz), 7.73 (1H, d, J=9.4 Hz), 7.34 (1H, d, J=9.3 Hz), 7.17 (1H, s),
6.14 (1H, s), 4.64 (2H, t, J=6.5 Hz), 4.11 (3H, s), 1.78 (2H, m),
1.45 (2H, m), 0.93 (3H, m). .sup.13C NMR (DMSO-d.sub.6, 1:4)
.delta. 185.2, 177.2, 175.4, 171.5, 155.8, 152.5, 137.4, 132.8,
127.1, 124.5, 123.0, 119.4, 111.5, 95.6, 70.8, 37.8, 31.7, 18.2,
13.5; FAB-MS: m/z=325.140 (calcd 325.139 for
C.sub.19H.sub.19NO.sub.4).
[0047] The physiochemical properties of compound of formula 1,
(wherein, X=OH, Y=Br, and Z=H, R.sup.2=CH.sub.3, 80-90%), mp
182-184.degree. C., IR (KBr) .nu..sub.max 3421, 2950, 1760, 1589
cm.sup.-1. .sup.1H-NMR (CDCl.sub.3+DMSO-d.sub.6, 1:4) .delta. 9.31
(1H, d, J=9.2 Hz), 8.12 (1H, s), 7.88 (1H, d, J=9.2 Hz), 7.3 (1H,
s), 6.15 (1H, s), 4.65 (2H, t, J=6.7 Hz), 4.09 (3H, s), 1.78 (2H,
m), 1.48 (2H, m), 0.96 (3H, m). .sup.13C NMR (DMSO-d.sub.6, 1:4)
.delta. 185.8, 177.3, 175.4, 172.0, 156.2, 136.2, 133.3, 126.1,
124.5, 123.2, 118.0, 111.4, 97.8, 71.8, 38.8, 31.5, 18.1, 13.3;
FAB-MS: m/z=404.256 (calcd 404.255 for
C.sub.19H.sub.18BrNO.sub.4).
[0048] The physiochemical properties of compound of the general
formula 1, (wherein, X=OCH.sub.2CH.sub.3, Y=H, and Z=H,
R.sup.2=CH.sub.3, 80-90%), mp 184-186.degree. C., IR (KBr)
.nu..sub.max 3016, 2923, 1756, 1601 cm.sup.-1. .sup.1H-NMR
(CDCl.sub.3) .delta. 9.41 (1H, d, J=9.3 Hz), 7.84 (1H, d, J=9.3
Hz), 7.63 (1H, d, J=9.4 Hz), 7.33 (1H, d, J=9.5 Hz), 7.09 (1H, s),
6.15 (1H, s), 4.71 (2H, t, J=6.6 Hz), 4.14 (2H, t, J=6.9 Hz), 4.05
(3H, s), 1.78 (5H, m), 1.49 (2H, m), 0.96 (3H, m). .sup.13C NMR
(CDCl.sub.3) .delta. 184.1, 176.3, 174.3, 170.2, 156.6, 153.3,
136.7, 126.6, 125.6, 122.8, 117.4, 109.5, 94.6, 71.9, 63.9, 37.1,
31.8, 18.2, 14.2, 13.5, 13.3; FAB-MS: m/z=353.164 (calcd 353.163
for C.sub.21H.sub.23NO.sub.4).
[0049] The physiochemical properties of compound of the general
formula 1, (wherein, X=C.sub.6H.sub.11O.sub.6, Y=H, Z=H,
R.sup.2=CH.sub.3, 85-90%), mp 162-164.degree. C.); IR (KBr)
.nu..sub.max 3029, 2943, 1762, 1591 cm.sup.-1; .sup.1H-NMR
(DMSO-d.sub.6) .delta. 9.19 (1H, d, J=9.4 Hz), 7.91 (1H, d, J=9.2
Hz), 7.73 (1H, d, J=9.4 Hz), 7.34 (1H, d, J=9.1 Hz), 7.17 (1H, s),
6.14 (1H, s), 6.01-5.62 (3H, m), 4.82-4.67 (2H, m), 3.98 (3H, s);
.sup.13C NMR (DMSO-d.sub.6) .delta. 197.72, 190.81, 190.27, 165.65,
164.15, 164.12, 162.76, 138.63, 135.59, 129.19, 129.11, 128.96,
128.95, 128.42, 121.93, 118.49, 117.39, 115.66, 102.44, 96.78,
76.14, 73.33, 66.66, 62.22, 35.25, 31.75, 19.07, 13.77; FAB-MS:
m/z=487.190 (calcd 487.184 for C.sub.25H.sub.29NO.sub.3).
EXAMPLE 2
General Procedure for Preparation of Compounds Represented by the
General Formula 2
[0050] A mixture of the corresponding quinaldinium salt (1 mmol),
and squaric acid (0.5 mmol) and quinoline (0.5 mL) was refluxed in
a mixture of n-butanol and benzene (6-9 mL each, 1:1) with
azeotropic distillation of water for 24-36 h. The solvent was
distilled off under reduced pressure to obtain a residue, which was
chromatographed over silica gel. Elution of the column with a
mixture (1:9) of methanol and chloroform gave the corresponding
squaraine dyes of the general formula 2.
[0051] The physiochemical properties of compound of the general
formula 2, (wherein, X=Br, Y=H, R.sup.2=CH.sub.3, 90-95%), mp
336-338.degree. C., IR (KBr) .nu..sub.max 3056, 1613, 1580
cm.sup.-1. .sup.1H-NMR (DMSO-d.sub.6) .delta. 9.31 (1H, d, J=9.5
Hz), 9.28 (1H, d, J=9.3 Hz), 7.3-8.4 (8H, m, aromatic), 5.86 (1H,
s, vinylic), 5.78(1H, s, vinylic), 3.85 (3H, s); 3.78 (3H, s);
FAB-MS: m/z=550.246 (calcd 550.248 for
C.sub.26H.sub.18Br.sub.2N.sub.2O.sub.2).
[0052] The physiochemical properties of compound of the general
formula 2, (wherein, X=I, Y=H, R.sup.2=CH.sub.3, 85-95%), mp
314-315.degree. C., IR (KBr) .nu..sub.max 2962, 1620, 1553
cm.sup.-1. .sup.1H-NMR (DMSO-d.sub.6) .delta. 9.23 (1H, d, J=9.4
Hz), 9.19 (1H, d, J=9.2 Hz), 7.1-8.05 (8H, m, aromatic), 5.89 (1H,
s, vinylic), 5.72 (1H, s, vinylic), 3.81 (3H, s), 3.71 (3H, s);
FAB-MS: m/z=643.944 (calcd 643.946 for
C.sub.26H.sub.18I.sub.2N.sub.2O.sub.2).
[0053] The physiochemical properties of compound of the general
formula 2, (wherein X=CN, Y=H, R.sup.2=CH.sub.3, 80-90%), mp
290-292.degree. C., IR (KBr) .nu..sub.max 3032, 1616, 1579
cm.sup.-1. .sup.1H-NMR (DMSO-d.sub.6) .delta. 9.49 (1H, d, J=9.4
Hz), 9.45 (1H, d, J=9.5 Hz), 7.4-8.27 (8H, m, aromatic), 5.92 (1H,
s, vinylic), 5.85 (1H, s, vinylic), 3.92 (3H, s), 3.84 (3H, s);
FAB-MS: m/z=498.149 (calcd 498.150 for
C.sub.27H.sub.22N.sub.4O.sub.6).
[0054] The physiochemical properties of compound of the general
formula 2, (wherein X=NO.sub.2, Y=H, R.sup.2=CH.sub.3, 75-82%), mp
306-308.degree. C., IR .nu..sub.max (KBr) 3029, 2225, 1602
cm.sup.-1; .sup.1H-NMR (DMSO-d.sub.6) .delta. 9.39 (1H, d, J=8.9
Hz), 9.42 (1H, d, J=9.4 Hz), 7.35-8.28 (8H, m, aromatic), 5.87 (1H,
s, vinylic), 5.79 (1H, s, vinylic), 3.95 (3H, s), 3.82 (3H, s);
FAB-MS: m/z=442.472 (calcd 442.470 for
C.sub.28H.sub.18Br.sub.2N.sub.4O.sub.2).
[0055] The physiochemical properties of compound of the general
formula 2, (wherein X=C.sub.27H.sub.43O.sub.3, (cholesterol), Y=H,
R.sup.2=CH.sub.3, 75-85%), mp 328-330.degree. C., .sup.1H-NMR
(CDCl.sub.3) .delta. 9.31(1H, d, J=9.57 Hz), 7.44-7.30 (4H, m,
aromatic), 5.80 (1H, s, vinylic), 5.43 (1H, s), 4.60 (1H, s), 3.79
(3H, s), 2.50 (3H, s); 2.04-0.69 (46H, m); FAB-MS: m/z=1204.821
(calcd 1204.815 for C.sub.80H.sub.104N.sub.2O.sub.7).
[0056] The physiochemical properties of the compound of the general
formula 2, (wherein X=C.sub.41H.sub.32O.sub.11 (.beta.-D-glucose
with the hydroxyl groups protected with Benzoyl chloride), Y=H,
R.sup.2=CH.sub.3, 75-80%), mp 294-296.degree. C., .sup.1H-NMR
(CDCl.sub.3) .delta. 9.17(1H, d, J=9.32 Hz), 7.98-7.86 (23H, m,
aromatic), 7.53-7.02 (47H, m, aromatic), 7.44-7.30 (4H, m,
aromatic), 6.01 (3H, m), 5.79-5.73 (7H, m), 5.46 (1H, s), 4.67-4.59
(5H, m), 4.17 (2H. s), 3.62 (6H, s); FAB-MS: m/z=1564.469 (calcd
1564.478 for C.sub.94H.sub.72N.sub.2O.sub.21).
EXAMPLE 3
General Procedure for Preparation of Compounds Represented by the
General Formula 3
[0057] A mixture of the corresponding semisquaraine (1 mmol) and
the iodo or bromo substituted quinaldinium salt (1 mmol), and
quinoline (0.5 mL) were refluxed in a mixture of n-butanol and
benzene (6-9 mL each, 1:1) with azeotropic distillation of water
for 18-24 h. The solvent was distilled off under reduced pressure
to obtain a residue, which was chromatographed over silica gel.
Elution of the column with a mixture (1:9) of methanol and
chloroform gave the corresponding squaraine dye.
[0058] The physiochemical properties of compound of the general
formula 3, (wherein X=OH, Q=I, Y=H, Z=H, R.sup.2=CH.sub.3, 90%),
325.degree. C., IR (KBr) .nu..sub.max 3442, 3046, 1729, 1621, 1579,
1560 cm.sup.-1. .sup.1H-NMR (DMSO-d.sub.6) .delta. 10.2 (1H, OH),
9.29 (1H, d, J=9.2 Hz), 9.01 (1H, d, J=9.5 Hz), 7.86-7.01 (8H, m),
5.77 (1H, s, vinylic), 5.44 (1H, s, vinylic), 3.9 (3H, s), 3.5 (3H,
s); FAB-MS: m/z=534.040 (calcd 534.039 for
C.sub.26H.sub.19IN.sub.2O.sub.3).
[0059] The physiochemical properties of compound of the general
formula 3, (wherein X=C.sub.6H.sub.11O.sub.6 (.beta.-D-glucose),
Q=I, Y=H, Z=H, R.sup.2=CH.sub.3, 85-92%), mp 316-318.degree. C.,
.sup.1H-NMR (DMSO-d.sub.6) .delta. 9.32 (1H, d, J=9.2 Hz), 9.13
(1H, d, J=9.5 Hz), 8.10-7.21 (8H, m), 6.12-6.01 (3H, m), 5.82-5.68
(4H, m), 3.86 (3H, s), 3.76 (3H, s); FAB-MS: m/z=696.091 (calcd
696.097 for C.sub.32H.sub.29IN.sub.2O.sub.8).
[0060] The physiochemical properties of compound of the general
formula 3, (wherein, X=C.sub.41H.sub.32O.sub.11 (.beta.-D-glucose
with the hydroxyl groups protected with benzoyl chloride), Y=H,
Z=H, Q=I, R.sup.2=CH.sub.3, 85-95%), mp 302-304.degree. C.,
.sup.1H-NMR (CDCl.sub.3) .delta. 9.11 (1H, d, J=8.6 Hz), 7.98-7.86
(12H, m), 7.53-7.02 (13H, m), 6.01 (3H, m), 5.79-5.46 (4H, m), 5.46
(1H, d, 7.2 Hz), 3.9 (3H, s), 3.6 (3H, s); FAB-MS: m/z=1112.210
(calcd 1112.202 for C.sub.60H.sub.45IN.sub.2O.sub.12).
[0061] The physiochemical properties of compound of the general
formula 3, (wherein X=C.sub.27H.sub.43O.sub.3, cholesterol, Y=H,
Q=I, Z=H, R.sup.2=CH.sub.3, 86-90%), mp 328-330.degree. C.,
.sup.1H-NMR (CDCl.sub.3) .delta. 9.16 (1H, d, J=9.3 Hz), 9.08 (1H,
d, J=9.1 Hz), 8.46-7.81 (8H, m, aromatic), 5.93 (1H, s, vinylic),
5.84 (1H, s, vinylic), 5.4 (1H, s), 4.6 (1H, m), 3.94 (3H, s); 3.81
(3H, s), 2.04-0.69 (48H, m) FAB-MS: m/z=933.389 (calcd 933.382 for
C.sub.53H.sub.61IN.sub.2O.sub.5).
EXAMPLE 4
[0062] Semisquaraine of the general formula 1 in which a quinaldine
moiety is linked to the squaric acid through methylene bridge have
absorption around 480 nm with high molar extinction coefficient
(4.0.times.10.sup.4 M.sup.-1 cm.sup.-1) which depends on the
substitution on the quanaldine unit. For example semisquaraines
with --OH groups at the 6.sup.th position showed absorption maxima
at 477 nm while semisquaraines with --OEt group at 6.sup.th
position showed absorption maxima at 478 nm in methanol. They
showed good fluorescence emission with emission maximum ranging
from 550-600 nm. The absorption and fluorescence emission spectra
of representative semisquaraine of the general formula 1 where
X=OH, Y=H, Z=H, R.sup.2=CH.sub.3 are shown in FIG. 1.
EXAMPLE 5
[0063] Squaraine dyes of the general formula 2 showed good
absorption in the long wavelength region. Similar to the
semisquaraines the absorption spectra of these dyes were also
dependent on the substitution on the quinaldine moiety. For
example, dye of the general formula 2, where X=I, Y=H,
R.sup.2=CH.sub.3, showed absorption maxima at around 750 nm in
DMSO, whereas dyes of the general formula 2, wherein X=CN, Y=H,
R.sup.2=CH.sub.3 and X=NO.sub.2, Y=H, R.sup.2=CH.sub.3 showed
absorption maxima around 615 nm and 618 nm respectively, in
methanol. The presence of electron withdrawing groups in the
quinaldine ring shifts the absorption maxima towards the blue
region. Similarly the emission spectra of these dyes showed a shift
in the emission maxima depending on the substituents present. For
example, dye of the general formula 2, where X=I, Y=H,
R.sup.2=CH.sub.3, showed emission maxima at around 772 nm in
dimethyl sulfoxide, whereas dye of the general formula 2, wherein
X=CN, Y=H, R.sup.2=CH.sub.3 showed emission maxima around 647 nm,
in methanol. The absorption and fluorescence emission spectra of
representative symmetrical squaraine dye of the general formula 2,
where X=I, Y=H, R.sup.2=CH.sub.3 are shown in FIG. 2.
EXAMPLE 6
Demonstration of Formation of Unsymmetrical Squaraine Dyes of the
General Formula 3
[0064] FIG. 3 shows the absorption spectra recorded at various time
intervals for the reaction between semisquaraines of the general
formula 1, where X=OH, Y=H, Z=H, R.sup.2=CH.sub.3, and halogenated
(Iodo or Bromo) quinaldinium salts. As seen from the figure,
initially only the absorption from the semisquaraine is observed at
485 nm. As the reaction progresses a new band at 715 nm appears and
intensifies with time showing the formation of the dye.
Interestingly, it was observed that the semisquaraines of the
general formula 1 reacted only with quinaldinium salts having
electron withdrawing substituents like nitro, cyano and
electronegative groups like iodo and bromo. Reaction of the
semisquaraines with quinaldinium salts having electron donating
substituents like OH, OEt did not yield any products (Inset of FIG.
3). Thus the specificity of the semisquaraines towards quinaldinium
salts with electron withdrawing substituents is demonstrated, which
enables the synthesis of a library of unsymmetrical squaraine dyes
that can be used for photodynamic and therapeutical
application.
EXAMPLE 7
[0065] Similar to the symmetrical dyes, unsymmetrical squaraine
dyes of the general formula 3 showed good absorption in the long
wavelength region. For example dye of the general formulae 3,
wherein X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3,
and X=OH, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3 showed absorption maxima
in the range 745-755 nm and emission maxima in the range 765-775 nm
in DMSO. The absorption and fluorescence emission spectra of
representative unsymmetrical squaraine dyes of the general formula
3, where X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3,
are shown in FIG. 4.
EXAMPLE 8
Cytotoxicity Induced by Squaraine Dyes of the General Formulae 2
and 3
[0066] In order to analyze the cytotoxicity of the squaraine dyes
of the general formula 2 wherein X=I, Y=H, R.sup.2=CH.sub.3 and 3
wherein X=OH, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3, with and without
visible light, the cloning efficiencies of L1210 mouse lymphoma
cells exposed to various concentrations of the dyes were
determined. The light exposure was carried out from a 1000 W
halogen lamp at a distance of 33 cm in Ca.sup.2+ and Mg.sup.2+ free
PBS (140 mM NaCl, 3 mM KCl, 8 mM Na.sub.2PH.sub.4, pH 7.4) on ice
(10.sup.6 cells/mL). Illumination for 10 min corresponds to 225
kJ/m.sup.2 between 400 and 800 nm. The cells were pelleted by
centrifugation and resuspended in PBSG three times. The cells were
resuspended at 3.times.10.sup.4 cells/mL in fresh medium at
37.degree. C. and the numbers of cells were counted repeatedly for
60 h. From the exponential part of the growth curves (between 24
and 60 h) the number of proliferating cells at the time of
resuspension was calculated by extrapolation. Cell survival was
defined as the ratio between proliferating and resuspended cells.
Results obtained with representative quinaldine based squaraine
dyes of the general formula 2 wherein X=I, Y=H, R.sup.2=CH.sub.3
and 3 wherein X=OH, Y=H, Q=I, Z=H, are shown in FIG. 5.
[0067] From the results it is clear that the squaraine dyes of the
general formula 2 wherein X=I, Y=H, R.sup.2=CH.sub.3 and 3 wherein
X=OH, Y=H, Q=I, Z=H, R.sup.2=CH.sub.3, show a dose dependent
photocytotoxicity. i.e. the % of cell survival decreases with the
increasing concentration of dyes of the formula the general formula
2 and 3 under illumination conditions indicating their high cell
killing efficiency under these conditions. At the same time these
dyes are found to be less toxic in the dark indicating their
non-toxicity in the absence of light. These results clearly
demonstrate the photodynamic therapeutical applications of
quinaldine based squaraine dyes. Of the two, dye of the general
formula 2, wherein X=I, Y=H, R.sup.2=CH.sub.3 is found to be more
toxic than the dye of the general formula 3, wherein X=OH, Y=H,
Q=I, Z=H, R.sup.2=CH.sub.3.
EXAMPLE 9
Cytotoxicity Induced by Squaraine Dyes of the General Formulae 3
where the Dyes are Conjugated with Cellular Recognition
Elements
[0068] The cytotoxicity induced by squaraine dyes of the general
formula 3 wherein X=C.sub.6H .sub.11O.sub.6, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 and X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 in mouse lymphoma cells L1210 with and without
visible light, was carried out as explained in EXAMPLE 8. Results
obtained with representative quinaldine based squaraine dyes of the
general formula 3 wherein X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 and X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 are shown in FIG. 6.
[0069] It is clear from FIG. 6 that the squaraine dyes of the
general formula 3 wherein X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 and X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 show a dose dependent photocytotoxicity. i.e. the
% of cell survival decreases with the increasing concentration of
dyes under illumination conditions indicating their high cell
killing efficiency under these conditions. These dyes show very
less toxicity in the absence of light thereby demonstrating their
photodynamic therapeutical application. The dye of the general
formula 3 wherein X=C.sub.6H.sub.11O.sub.6, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3 showed higher toxicity than the dye of the general
formula 3 wherein X=C.sub.27H.sub.43O.sub.3, Y=H, Q=I, Z=H,
R.sup.2=CH.sub.3.
[0070] The quinaldine based squaraine dyes of the present invention
possess satisfactory properties of a photosensitizer for
photodynamic therapeutical and industrial applications.
The Main Advantages of these Systems Include:
[0071] 1. Semisquaraines, symmetrical and unsymmetrical squaraine
dyes represented by formulae 1, 2 and 3 are novel and pure single
substances. [0072] 2. Their synthetic methodology is very
economical [0073] 3. Symmetrical and unsymmetrical squaraine dyes
represented by formulae 2 and 3 possess absorption in the
near-infrared region (700-750 nm) [0074] 4. They are nontoxic in
the dark but show good cell killing properties when exposed to
light, as expected of an ideal photosensitizer [0075] 5. Their
benzo ring can be substituted with various functional groups so as
to have increased cellular uptake and localisation. [0076] 6. They
can be conjugated with cellular recognition elements like
cholesterol and sugar so as to bring about site specific tumor
localisation and destruction. [0077] 7. They can be used for
photodynamic industrial applications such as sterilization of water
etc. [0078] 8. They can be used as near-infrared fluorescence
sensors in biological applications. [0079] 9. They can be used for
the detection of biologically important metal ions under
physiological conditions.
* * * * *